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From 2010.igem.org

Materials

• Pipettes
• Universal tubes
• Streaking loop
• Christensen's Urea Agar
  • Make up 1 liter of the agar mixture containing:
  1. Gelatine peptone 1.0g
  2. Potassium dihydrogen phosphate 2.0g
  3. Sodium chloride 5.0g
  4. Agar 20g
  • Top up to 1 liter and apply gentle heating to dissolve. Sterilize at 115°C for at least 20min.
  • Add the following to the molten base:
  1. D(+)-Glucose 1.0g
  2. Phenol red, 0.2% 6ml
  • Note: Ensure that the work was done using aseptic technique.
  • Transfer 10ml of the molten base to universal tube and allow the agar to set in a slanted position.
  • Store the harden agar in the fridge.

Protocol

1. Perform the experiment using aseptic technique.
2. Pick up single colony of B. subtilis 168 and streaking it onto the slanted agar tube.
3. Loosen up the cap to allow air exchange between the interior of the tube and the external environment.
4. Incubate the tubes overnight at 37°C.
5. Set up the tubes as indicated:
   1. Negative control - Without B. subtilis 168
   2. Test 1 (Duplicate) - Inoculated with B. subtilis 168
   3. Test 2 (Duplicate) - Inoculated with B. subtilis 168